The generation of free radicals and toxic lipoperoxides is an important component of melanogenesis and phagocytosis by other non-retinal cell types but has not been investigated in the retinal pigment epithelium (RPE). Since the viability of aerobic cells is dependent on the ability to detoxify lipoperoxides, usually with glutathione peroxidase, it is reasonable to hypothesize that melanogenesis and phagocytosis by RPE involves the removal of peroxides too. When glutathione peroxidase is deficient, neutrophils cease to phagocytose, suggesting autoregulation sensitive to a component of the peroxidase redox system. We have determined that porcine RPE in culture maintain differentiated characteristics including the continued production of tyrosinase. We will use this system to study: 1) Production of lipoperoxides during melanogenesis and phagocytosis: We will determine whether phagocytosis is associated with a burst of peroxides as it is in macrophages and neutrophils, and what is the effect of type and quantity of material phagocytoses and what is the relationship between melanogenesis and lipoperoxide. For these studies we will assay glutathione and p-phenylominediamine peroxidases, reduced and oxidized glutathione and NADP, lipoperoxide production and lipofuscin content (as an index of cumulative peroxidative damage). 2) The influence of peroxidation on phagocytosis and melanogenesis. We will experimentally manipulate the level of intracellular peroxides and components of the peroxidase redox system and study the effect on melanogenic and phagocytic capacity. Similarly, we'll determine whether control and melanogenic cells have equal capacity to phagocytose, and vice versa. 3) Regulation of melanogenesis: We will use the most effective of several possible melanogenic stimuli to determine whether they operate by altering cellular inhibitors, and whether concurrent stimulation of the peroxidase system elevates melanogenesis. For this we will assay tyrosinase, concentration of premelanogens, melanin and lipofuscin.